Thermotoga neapolitana bgIB gene, upstream of lamA, encodes a highly thermostable β-glucosidase that is a laminaribiase

The gene for thermostable 1,3-beta-glucosidase BglB was cloned from the chromosome of Thermotoga neapolitana and its primary sequence was determined. The purified recombinant beta-glucosidase B had a monomer molecular mass of 81 kDa in accordance with the amino acid sequence predicted from the nucleotide sequence of clone pTT51. It was a member of glycosylhydrolase family 3 and belonged to enzyme class EC 3.2.1.21. beta-Glucosidase B had a specific activity of 255 U mg-1 on 4-nitrophenyl(PNP)-beta-glucoside at the optima of pH (5.5) and temperature (90 degrees C), and K(m) values of 0.1, 10 and 50 mM for PNP-beta-glucoside, laminaribiose and cellobiose, respectively. The gene bglB was located immediately upstream of the laminarinase gene lamA. Both genes were transcribed from the same DNA strand and were not separated by a palindromic transcription terminator. The two purified enzymes 1,3-beta-glucosidase BglB (laminaribiase) and 1,3-beta-glucanase LamA (laminarinase) were together capable of completely degrading laminarin to glucose.